The ANPEP Knockout HT29 Polyclonal Cells product is a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 colorectal adenocarcinoma cell line. This model features disruption of the ANPEP gene, encoding aminopeptidase N (CD13), resulting in a loss-of-function system suitable for investigating CD13-dependent processes. The polyclonal nature of the knockout pool provides a heterogeneous population of gene-edited cells, avoiding clonal artifacts and capturing a range of editing outcomes without selection for a single clone. Validated for absence of CD13 expression through standard molecular biology techniques, this product serves as a robust tool for functional genomics and cancer biology studies.
HT29 cells are a well-characterized human colorectal adenocarcinoma line originating from a primary tumor, capable of differentiating into enterocyte-like cells upon reaching confluence or in response to sodium butyrate. They exhibit a mucin-secreting phenotype and are widely used as an intestinal epithelial model to study colorectal cancer, mucosal barrier function, and differentiation pathways. The HT29 background provides a physiologically relevant context for examining the role of CD13 in intestinal epithelial biology, as the line retains key features of colorectal tumorigenesis and epithelial-mesenchymal plasticity.
The ANPEP gene product, CD13, is a membrane-bound zinc metalloprotease that preferentially cleaves neutral amino acids from the N-terminus of peptides, including enkephalins and angiotensin. It is regulated by cytokines such as TNF-alpha, IL-4, and IL-13, as well as retinoic acid and C/EBP transcription factors. CD13 directly interacts with integrin beta-1, galectin-3, and extracellular matrix components like collagen IV and fibronectin, forming focal adhesion complexes that activate focal adhesion kinase (FAK) and downstream ERK/MAPK signaling. This network transduces signals from the extracellular matrix to control cell adhesion, proliferation, migration, and angiogenesis. CD13 also processes chemokines and other peptides, modulating immune cell recruitment and the tumor microenvironment. Disruption of ANPEP therefore abrogates these molecular interactions and signaling cascades.
In the HT29 colorectal adenocarcinoma context, ANPEP knockout eliminates aminopeptidase N activity, leading to impaired integrin-mediated adhesion and reduced invasive potential. The loss of CD13 attenuates peptide cleavage that normally contributes to autocrine growth stimuli and matrix remodeling. This recapitulates key aspects of therapeutic CD13 targeting, making the model valuable for preclinical drug validation. Moreover, HT29 cells?? capacity to undergo enterocytic differentiation allows researchers to explore how CD13 loss influences barrier integrity, lumen formation, and mucin production in a colorectal cancer setting, linking to pathologies such as inflammatory bowel disease and metastatic progression.
Researchers can employ this polyclonal knockout model in diverse experimental workflows, including tumor microenvironment remodeling studies, angiogenesis assays (e.g., tube formation), and drug target validation. Representative assays encompass western blotting for CD13, enzymatic activity measurements, MTT proliferation assays, Transwell migration/invasion assays, flow cytometry for surface CD13, RT-qPCR for ANPEP mRNA, and immunofluorescence for CD13 localization. These applications enable detailed dissection of CD13??s role in integrin signaling, peptide hormone regulation, and intestinal barrier function. For further information or to discuss custom applications, please contact Ascent Research.
